Numerous compounds are currently undergoing in vitro development and clinical evaluation as potential drugs for the treatment of human immunodeficiency virus type 1(HIV-1) infection and the associated acquired immunodeficiency syndrome (AIDS). Historically, compounds directed toward inhibition of virus attachment to target cells have failed clinically due to their toxicities at effective antiviral concentrations, poor absorption from the gut or lack of broad spectrum activities against clinical stains of HIV-1. Similarly, the approach to use synthetic nucleoside analogs, such as 3'-azido-3'-deoxythymidine (AZT), or the complex class of non-nucleoside compounds to target the HIV-1 reverse transcriptase (RT) enzyme has been plagued by the emergence of drug-resistant strains. HIV-1 protease has also become the focus of much attention as a potential antiviral target due to its critical role in the post-integration processing of viral precursor polypeptides to their mature products, a process required for maturation of virus particles into infectious virions. Unfortunately, the vast majority of designed inhibitors of protease are substrate-based peptide structures that typically demonstrate poor bioavailability, short serum half-lives and overt cytotoxicity at effective antiviral concentration.
In addition, most antiviral drugs used to control the spread of HIV-1 have also proven to become compromised under the selection pressure of the drug, as the virus soon mutates to a drug-resistant strain. This tendency to develop drug resistance is a survival strategy used by many classes of viruses and is particularly pronounced among the members of the retrovirus family. One way to defeat this survival strategy is to focus on drugs attacking specific elements of the virus that are intolerant to mutations. Such elements can be identified by searching the proteins present in all viruses within the virus class to identify common or highly conserved structures.
Retroviruses have a highly conserved structure in their nucleocapsid (NC) proteins. All NC proteins of the Oncoviridae and Lentiviridae subfamilies of Retroviridae contain sequences of 14 amino acids with 4 invariant residues, Cys(X).sub.2 Cys(X).sub.4 His(X).sub.4 Cys, which chelate zinc through histidine imidazole and cysteine thiolates with a K.sub.d less than 10.sup.-13. These structures are referred to as retroviral CCHC zinc fingers, and are one of the most highly conserved features of retroviruses (Henderson, et al., J. Biol. Chem. 256:8400-8406 (1981)). Examples of retroviruses which possess at least one CCHC type zinc finger per nucleocapsid protein include, but are not limited to, HIV-1, HIV-2, SIV, BIV, EIAV, Visna, CaEV, HTLV-1, BLV, MPMV, MMTV, RSV, MuLV, FeLV, BaEV, and SSV. Due to their highly conserved nature, it is thought that CCHC zinc fingers perform an essential function in viral infectivity. In fact, it has been disclosed that mutations of the chelating residues (CCHC) in the zinc fingers yield a non-infectious virus (Gorelick, et al., J. Virol. 64:3207-3211 (1990)).
HIV-1 NC contains two zinc finger domains separated by only 7 amino acids. HIV-1 NC proteins are synthesized as part of the Pr55.sup.gag and Pr160.sup.gag-pol precursor polyproteins, and the fingers within these precursor molecules are required for packaging of viral genomic RNA and to form the core structure of the immature virion. Subsequent proteolytic processing of these precursors yields the mature p7NC protein, and the fingers of the NC protein are required for the virus to fully execute reverse transcription in the next target cell. Hence, treating of HIV-1 infected individuals with antiviral compounds that target the mutationally intolerant retroviral zinc finger may provide for multiple inhibitory effects on the viral replication cycle while attenuating the emergence of drug-resistant HIV-1 strains.
Widespread acceptance of the CCHC zinc finger as an antiviral target has not been forthcoming due to a lack of identification of compounds that selectively target that structure. Recently, however, it has been demonstrated that the two CCHC zinc fingers of the HIV-1 p7NC protein are susceptible to electrophilic attack by certain electrophilic reagents (nitrosos, disulfides, disulfoxides, maleamides, peroxides, and others), resulting in covalent modification of the Cys sulfur atoms and functional inactivation of the fingers.
For example, it has been shown that the CCHC zinc fingers could be specifically attacked by the thiolate-reactive C-nitroso compounds, resulting in inactivation of HIV-1 and SIV infectivity (Rice et al., Nature 361:473-475 (1993)).
The action of the electrophilic C-nitroso compounds is through a chemical attack of the compound on the nucleophilic zinc-coordinating cysteine thiolates, with subsequent ejection of zinc from the structure; the action is not mediated by a chelation effect.
In addition to the C-nitroso compounds, a second class of compounds has been found which targets the zinc finger of retroviruses. This second class of compounds falls into the general class of disulfide benzamides (DIBAs) (Rice et al., Science 270: 1194-1197, (1995)). It has been shown that the DIBAs are capable of inhibiting retroviruses. The compounds do not affect virus binding to cells or the activities of purified HIV-1 reverse transcriptase or integrase, and protease inhibition does not correlate with antiviral activity in culture. The DIBAs directly inactivate HIV- 1 virions by entering the virions and cross-linking the p7NC proteins. In addition, DIBAs inhibit the production of infectious virus from previously infected cells by acting on the zinc fingers in the Gag precursor polyproteins. The compounds are also synergistic with other antiviral agents and drug-resistant mutants have not arisen. Moreover, the DIBA compounds do not affect the activity of proteins tested to date that contain the classical type CCCC or CCHH zinc finger motifs.
Despite the promising antiviral activity of the DIBA type compounds, there is the possibility that in vivo the sulfur atoms in these compounds could be reduced. The resultant two inactive monomers could disassociate resulting in a loss of antiviral activity.